Brucellosis is a zoonotic infection mainly affecting farm animals
such as cattle, sheep and goats. Brucella species primarily infect the
reproductive tract, causing spontaneous abortions and infertility in
livestock. Brucellosis has a worldwide distribution, but is most
prevalent in areas with poorly established domestic animal and public
health programmes. [1,2] According to the US Centers for Disease Control
and Prevention, Africa is among the areas most commonly affected.

Human brucellosis is thought to be considerably under-diagnosed,
and although it is a notifiable disease in South Africa (SA), as in many
parts of the world, it is probably under-reported. [1] Fig. 1 indicates
the estimated seroprevalence of B. abortus in animals in SA. No formal
B. melitensis surveillance occurs in SA.

Humans are infected with Brucella by contact with sick animals or
infected animal products. Organisms can be inoculated via cuts and/ or
abrasions when handling infected animal carcasses or products of
conception, or inhaled as aerosols during these and other procedures.
Ingestion of unpasteurised milk/dairy products or undercooked meat can
also lead to infection. High-risk individuals include farmers,
veterinarians, abattoir workers and meat handlers. [2] Laboratory
workers are also at risk if inadvertently exposed to live cultures of
Brucella.

Most cases of human brucellosis are caused by B. melitensis, which
is the species associated with more severe disease. The usual animal
reservoirs for Brucella are sheep and goats (B. melitensis), cattle (B.
abortus), and pigs (B. suis). Disease in humans may have an acute or
subacute onset, with the illness having a propensity to become chronic
and relapsing. Brucellosis is a systemic disease, usually presenting as
a febrile illness with constitutional symptoms such as anorexia,
malaise, headaches and chills. The patient may complain of arthralgia
and back pain, as well as abdominal pain. Clinical findings include
lymphadenopathy and hepatosplenomegaly. Up to 40% of patients with
brucellosis have osteoarticular complications. [2] Chronic brucellosis
is difficult to treat, requiring prolonged antibiotics and in some cases
surgery.

A definitive diagnosis of Brucella infection is established by
isolating the organism from blood, bone marrow, cerebrospinal fluid,
tissue, pus or other relevant samples. [2] A presumptive identification
of the cultured isolate can be made using basic biochemical tests and
colonial morphology. Cultured isolates should be referred to a
specialised facility for further confirmatory testing and speciation,
which can be done using molecular methods such as the Brucella-specific,
Bruce-ladder multiplex polymerase chain reaction (PCR) and biotyping.
[1,3,4]

Case report

A 27-year-old man from a town in the Great Karoo, Western Cape
Province, SA, initially presented to a local district hospital in
November 2014 with a history of lower back pain and fever with new onset
of vomiting and diarrhoea. On examination he appeared chronically unwell
and had abdominal tenderness. Initial blood results showed pancytopenia,
a normocytic, normochromic anaemia, and mildly elevated transaminases
and canalicular enzymes. An abdominal ultrasound scan revealed diffuse
hepatosplenomegaly with no granulomas or other focal lesions. One of two
blood cultures sent by the regional hospital was positive for
Gram-negative bacilli after 3 days of incubation. This provisional
result was communicated to the treating clinician, who commenced
ceftriaxone empirically. The bone marrow report showed prominent serous
atrophy and listed common causes for this, which included chronic
illnesses such as carcinoma, tuberculosis and other chronic infections.
No granulomas were observed. On provisional testing of the cultured
isolate, Brucella was suspected and the national and provincial
departments of health were notified, prompting a veterinary
investigation. An in-depth history revealed that the patient had
regularly fed his dog with cattle, sheep and goat abattoir waste that
was disposed of at an open-access municipal waste site in the local
town. There was no history of consumption of unpasteurised milk.

The cultured isolate was subsequently confirmed to be B. melitensis
by the Department of Veterinary Tropical Disease at the University of
Pretoria, using Bruce-ladder multiplex PCR. [3] Biotyping performed by
the Agricultural Research Council-Onderstepoort Veterinary Institute
identified the isolate as B. melitensis biovar 1. [4]

Laboratory exposure

By the time Brucella was suspected as the infecting agent, staff at
two medical microbiology laboratories involved had potentially been
exposed. Management of staff, which included post-exposure prophylaxis,
varied depending on the risk category. [5] The exposed laboratory
workers were followed up by occupational health and safety staff for 24
weeks.

A total of 75 laboratory staff/visitors were identified as having
been exposed to the organism. To make a serological diagnosis of
Brucella, paired sera are collected at least 2 - 4 weeks apart looking
for a four-fold or greater rise in antibody titre. [6] There are
numerous commercial kits and serological methods commonly used and
recommended, including the Rose Bengal test, serum tube agglutination
and enzyme-linked immunosorbent assay (ELISA). [2] Serological testing
using an ELISA, MASTAZYME BRUCELLA (MAST Diagnostics, UK), was performed
for all exposed staff members at baseline and then at 6 and 24 weeks.
[6]

Five staff members had borderline or low-level positive IgG or IgM
levels detected on serological testing at baseline. Repeat testing of
the same samples as well as repeat samples sent 3 - 4 weeks later
revealed considerable variability, particularly in IgM results.

In the absence of local studies to determine appropriate cut-off
values in SA, and in view of the known intrinsic variability of IgM
assays as well as known cross-reactivity with Enterobacteriaceae,
borderline results can be difficult to interpret. Failure to demonstrate
a (four-fold or greater) rise in the titre for specific antibody in
paired sera usually excludes recent infection. [6] None of the staff
members tested had significant increases in titre (four-fold or greater
rise in immunoglobulins) over the 24-week period. All exposed staff
members remained asymptomatic throughout the 6-month follow-up period.

Informed consent was obtained from the patient, and the study was
approved by the Human Research Ethics Committee, University of Cape
Town, SA (HREC REF: 220/2015).

Veterinary investigation

The state veterinarian's investigation revealed that two other
people who had lambed goats on a nearby farm in 2014 had become ill and
were subsequently diagnosed with brucellosis, based on serological
testing. The state veterinarian placed the farm under quarantine and
initiated sampling of the cattle, sheep and goats. Animals on
surrounding farms were also sampled. Serological testing using either
the Rose Bengal test or the complement fixation test was performed.
These methods detect nonspecific antibodies to both B. melitensis and B.
abortus.

Of the 100 goats sampled on the farm in question, 44 tested
positive for Brucella on serological testing. All animals on the farm
were quarantined, but none had been sacrificed at the time of writing
and no samples were available for culture and confirmation of the
Brucella species. As goats do not contract B. abortus, it was concluded
that the most likely strain causing the outbreak was B. melitensis. No
animals on other surrounding farms had tested positive for Brucella at
the time of writing.

Discussion

The Department of Agriculture, Forestry and Fisheries relies on
notifications of diseases and important epidemiological incidents from
veterinarians. The first documented cases of veterinary B. melitensis in
SA were in sheep in 1965 in the Transvaal Province. [7] Vaccination is
the foundation of a functional Brucella control programme in livestock.
This involves the B. abortus strain 19 (S19) and B. abortus RB51
vaccines for cattle and B. melitensis Rev. 1 vaccine for sheep and
goats. These are both live attenuated vaccines with the potential of
being pathogenic to humans if not administered appropriately. [1,8]
Vaccination regulations in SA as stipulated by the Animal Diseases Act
35 of 1984 have specific guidelines on vaccination of animals against
Brucella. There are currently no safe, efficacious vaccines recommended
for routine use in prevention of brucellosis in at-risk humans.
Prevention strategies therefore require robust animal programmes and
adherence to prevention/eradication and control protocols. [1] From a
public health perspective, the key targets would be the prevention of
infection due to direct/indirect contact with infected livestock or live
vaccines and prevention of food-borne illness, as well as strengthening
the diagnosis and management of human cases with increased awareness and
safe laboratory practices when Brucella species are suspected. From a
veterinary perspective, occupational hygiene involves safe injection
practices for animals, monitoring and surveillance of disease in
livestock, and having adequate protocols in place to manage an outbreak
effectively. The prevention of food-borne illness includes community
education campaigns around the consumption of unpasteurised dairy
products or undercooked meat.

[FIGURE 1 OMITTED]

Patients with brucellosis often present with nonspecific signs and
symptoms. Symptoms may persist for weeks to months. In the context of a
country with a high prevalence of tuberculosis and HIV infection,
patients presenting with features of chronic brucellosis may be
misdiagnosed and mistreated. As outbreaks of B. melitensis in animals in
SA are generally infrequent[9] and human cases are even less common,
there is a lack of awareness among clinicians and laboratory staff, who
have limited experience in diagnosing and handling this organism, with
subsequent delay in diagnosis. A high index of suspicion and a proper
social and exposure history are therefore essential in making a timely
diagnosis. Effective communication between the clinician and the
laboratory regarding suspected infections is imperative to ensure that
all biological samples are handled appropriately. The testing laboratory
should have protocols in place to ensure staff safety and maintain good
safety practices. When working with a World Health Organization
biosafety risk level 3 organism such as Brucella, all work should also
be performed in an appropriate biosafety cabinet, in a biosafety level 3
facility, by individuals trained for this work. [10]

Conclusion

A multidisciplinary Brucella control programme can be effective in
preventing human infections with an approach that integrates three key
elements: veterinary service, public health, and the medical healthcare
system.